胶孢炭疽菌T-DNA插入突变体库的构建及其分子分析  

Construction of a T-DNA Insertional Mutant Library of Colletotrichum gloeosporioides of Mango and Its Molecular Analysis

作　　者：黄贵修[1] 薛迎春[1] 卢昕[1] 

机构地区：[1]中国热带农业科学院环境与植物保护研究所海南省热带农业有害生物检测监控重点实验室,海南儋州571737

出　　处：《热带作物学报》2008年第1期27-32,共6页Chinese Journal of Tropical Crops

基　　金：海南省教育厅高等学校科研资助重点项目(Hjkj200321);中央级公益性科研院所基本科研业务费专项(No.2007hzs1J003);中国热带农业科学院科技基金项目(RKY0704)

摘　　要：利用根癌农杆菌介导的T-DNA插入遗传转化体系转化杧果炭疽菌SC2菌株,共获得抗潮霉素B突变体4680个,平均每转化1.0×106个炭疽菌分生孢子可得到300～980个突变体,且潮霉素抗性在多次传代实验中得到稳定遗传。随机挑取38个突变体进行PCR检测,均可扩增出1条约1.2kb的目标谱带,说明突变体为T-DNA插入引起;进一步的Southern blot杂交分析结果表明,在随机挑选的20个突变体中,有1个(5%)为三拷贝T-DNA插入,4个(20%)为双拷贝插入,剩余的15个(75%)为单拷贝插入。Mangifera anthracnose, caused by Colletotrichum gloeosporioides,is one of the most damaging phytopathogens of mango（Mangifera indica L） in the world.A highly efficient Agrobacterium tumefaciens-mediated transformation of C, gloeosporioides was established. Strain SC2 of C. gloeosporioides was mutated through T-DNA insertion mediated by A. tumefaciens strain Agl-1. The transformation efficiency ranged from a low of 300 to a high of 980 transformants/10^6 conidia, and 4 680 transformants were obtained. The PCR amplification found that 38 tested transformants randomly selected were all resistant to hygromycin B and contained hph gene, indicating that the transformants were T-DNA inserted mutants. Southern blotting analysis showed the 20 T-DNA inserted

关 键 词：胶孢炭疽菌 T-DNA 插入突变 PCR SOUTHERN杂交 

分 类 号：S667.7[农业科学—果树学] Q785[农业科学—园艺学]